Method for treating neurological disorders, including tumors, with electroporation

ABSTRACT

This disclosure describes the methods, devices, and systems of treating diseased tissue with integrated nanosecond pulse irreversible electroporation. Methods and systems as disclosed provide MRI compatible shielded electrodes and electrode leads to prevent emanating radiofrequency noise and improve image quality, disconnecting the electrode from the cable linkage to the pulse generator reduce electromagnetic interference and image artifacts, placing electrodes strategically within a guide cannula to minimize distortion from heterogeneities or maximize ablation within the tissue, utilizing conductive fluids, innate or external, such as cerebral spinal fluid or grounding pads to provide a pathway for current return, and for timing of the electrical waveforms with inherent brain electrical activity.

FIELD OF THE INVENTION

The present disclosure relates to the use of electroporation fortreatment of diseased tissue within the brain, head, neck, and spinalcord in relation to treatment of diseases and disorders. The presentdisclosure relates more specifically to devices and methods fordestroying aberrant cell masses, including tumor tissues, with use ofwaveforms capable of inducing electroporation. The disclosure alsorelates to procedures to enhance the safety and treatment outcomes ofelectroporation intervention. The present disclosure also relates toelectroporation with placement of electrodes to minimize distortion fromheterogeneities, maximize ablation within tissue, or both, such as byplacement of electrodes within a guide cannula. The disclosure alsorelates to electroporation with a function generator operable to deliverpulse parameters desired for such procedures, and a function generatorconfigurable for such operation.

BACKGROUND

The Central Brain Registry of the United States (CBTRUS) estimates that79,270 Americans may be diagnosed with primary malignant (26,070) andprimary non-malignant (53,200) brain and central nervous system (CNS)tumors in 2017. Additionally, it is estimated that 20-40% of all othercancers (˜250,000) including lung, breast, melanoma, colon, and kidneyeventually metastasize to the brain. Gliomas are primary tumors thatarise from glial cells that normally support and protect neurons in thebrain and the CNS. Gliomas represent about 80% of all primary malignantbrain tumors with an estimated annual incidence of 20,856 Americans in2017. Unfortunately, Glioblastoma Multiforme (GBM=grade IV astrocytoma)accounts for 55.1% (12,120 Americans) of all malignant gliomas (MG)making it the most common and deadliest brain tumor. Despite efforts todefeat malignant glioma, 16,947 Americans may die from primary malignantbrain and other CNS tumors in 2017. Thus, there is a dire need for newtechnologies that may alter the course of malignant brain tumortreatment and improve survival and quality-of-life of patients sufferingfrom this and other deadly diseases.

The incidence of aggressive brain malignancies is increasing, in partdue to the aging population in the US. The poor prognosis for MGpatients results from a lack of effective targeted therapies combinedwith the failure of chemotherapies to penetrate the diffuse tumorinfiltrative niche that causes recurrence. A treatment approach usingelectric field-based cell-selective tumor ablation may lead to moreeffective MG treatment. Primary brain tumors, such as MG, are commonlytreated through surgery, radiation therapy, and/or chemotherapeuticregimens. Even when using aggressive and often debilitating therapies,cure rates have not improved. Regrowth of the primary tumor is virtuallyinevitable. Nearly all patients with MG develop tumor resistance totreatment with rapid case progression. One of the main reasons for poorsurvival is migration and continued growth of glioma cells severalcentimeters beyond the margins of the observable tumor. These distant,infiltrating cells are a key factor in tumor progression and resistanceto therapy.

Irreversible electroporation (IRE) is a tumor treatment method fortissue ablation that does not require elevating the tumor to lethaltemperatures to achieve lasting therapeutic effects. An example isdisclosed in U.S. Pat. No. 8,992,517B2 to Davalos et al. The IREtechnique uses microsecond-long (˜100 μs) electric pulses, applied withminimally invasive electrodes in the targeted region to generateelectric fields in excess of 1000 V/cm and kill tissue. This ablationmodality is capable of killing cells in large volumes (>50 cm³) withoutinducing thermal damage to essential tissue components, such asextracellular matrix, blood vessels, and nerves from damage. Recently,IRE has been shown to nearly double the median survival of patientsdiagnosed with stage III pancreatic cancer (from 6-13 months to 24.9months).

IRE systems and methods using bipolar pulses also may be used fortreatment of neurological disorders. Such systems and methods aredisclosed in U.S. Pat. No. 8,926,606B2, U.S. Pat. No. 8,992,517, andUS20160338758. A suitable commercially available system, for example, isthe H-FIRE™ Integrated Nanosecond Pulsed IRE system available from thepresent applicant (Voltmed Inc., Blackburg, Va.). In such an IntegratedNanosecond Pulsed IRE system, the relatively long (˜100 μs) IREelectrical pulses are replaced with bursts of ultrashort (˜0.5-2 μs)bipolar pulses. This change improves procedural safety, as a minimallyinvasive procedure may be performed without neuroparalytic agents. Thereis a latency period between the delivery of a single monophasic pulseand the firing of an action potential. Delivering an alternatingpolarity pulse within this latency period (˜100 μs) may acceleratemembrane repolarization and inhibit action potential generation.Electroporation still occurs, as the time course for pore formation issignificantly faster (˜500 ns). An Integrated Nanosecond Pulsed IREsystem has been used to safely perform electroporation treatment withoutneuroparalytic agents on three (3) canine patients with brain tumors.

An Integrated Nanosecond Pulsed IRE system may be tuned to provideselective cancer cell ablation within the normal brain tissue thatsurrounds MG. Malignant cells may exhibit an enlargednucleus-to-cytoplasm ratio (NCR), and the Integrated Nanosecond PulsedIRE system may be tuned to provide a frequency of polarity changes tomaximize the transmembrane potential on the plasma membrane according tothis geometrical consideration. It is believed that close proximity ofthe nuclear membrane to the plasma membrane in malignant cells maypromote rapid charging of the plasma membrane when high-frequencywaveforms are applied, and that electroporation may be achieved at alower electric field compared to healthy cells. Given the poor prognosisof aggressive MG, new methods capable of destroying MG without damagingsurrounding critical vascular structures are desired.

BRIEF SUMMARY OF THE INVENTION

The present disclosure provides improved Integrated Nanosecond PulsedIRE systems and apparatus, and improved methods for performingIntegrated Nanosecond Pulsed IRE. In embodiments, for example, anIntegrated Nanosecond Pulsed IRE system may include electrodes which arecompatible and usable with simultaneous operation of MRI equipment. Suchan Integrated Nanosecond Pulsed IRE system, which is compatible andusable with simultaneous operation of MRI equipment, may providevisibility of the electrodes and may enable the procedure and placementof the electrodes in an accurate, precise manner. In embodiments,improved control of placement of electrodes may provide improvedelectroporation of target tissue and improved selectivity by avoidingundesired electroporation of surrounding, non-target tissue. With MRIguidance before and during the procedure, the treatment may be monitoredas well as changed during the procedure, if needed. In embodiments, sucha system may be configured to provide synchronization of the guidecannula, pulses, or both, with inherent brain electrical activity. Suchsynchronization may enable minimizing distortion from heterogeneities,may maximize ablation within the tissue, and may enable improvedaccuracy in placement of the electrodes, and thus may provide moreaccuracy in defining or differentiating the treatment area or targetarea of tissues from surrounding, non-target tissues. In embodiments,such a system may include capability for the electrode to bedisconnected from a cable linkage to a pulse generator with reduction ofelectromagnetic interference, image artifacts, or both. In embodiments,a method for performing Integrated Nanosecond Pulsed IRE may includedisconnecting the electrode from a cable linkage to a pulse generatorwith reduced or limited electromagnetic interference, image artifacts,or both.

The present disclosure provide a method of electroporation treatment ofdiseased tissue, with improved safety and with improved treatmentoutcomes of electroporation therapy. Embodiments may include a method ofmaking an electrode for an electroporation system, that is compatibleand usable with simultaneous operation of MRI equipment. In embodiments,a method of electroporation may include operating an electroporationsystem having an electrode compatible and usable with simultaneousoperation of MRI equipment, for administering electroporation therapy.In embodiments, such a method may include shielding the electrode leadsto prevent electromagnetic interference of the leads, the electrode, orboth, with MRI equipment in operation with the electroporation system.In embodiments, a method for electroporation may include operatingmanual switches, electronic switches, or both, to prevent or reduceemanating radiofrequency noise to MRI equipment, and to improve MRIimage quality. In embodiments, such a method may include utilizingtissue property measurements, drug delivery systems, or both, incombination with an MRI-compatible electroporation electrode, to enableplacement of one or plural electroporation electrodes in tissue atselected electrode location(s) within a guide cannula. In embodiments,such a method may include placement of one or plural electroporationelectrodes in tissue, at selected location(s) within a guide cannula, tominimize distortion from heterogeneities or maximize ablation within thetissue. In embodiments, such a method may utilize conductive fluids,innate or external, such as cerebral spinal fluid, or grounding pads, toprovide a pathway for current return, and for timing of the electricalwaveforms with inherent brain electrical activity to enhance safety andtreatment outcomes for a patient undergoing an electroporationprocedure. Methods of electroporation as disclosed herein may be usedfor treatment of any ailment(s) within the brain, head, neck and spinalcord that may be treated with electroporation. Ailments treated bymethods of electroporation may include, without limitation, cancer,disorders, tumors, essential tremors, and Parkinson's disease.

Methods of electroporation disclosed herein may provide electroporationwaveforms for treatment of cancers, brain disorders and tumors viatissue ablation in combination with immunotherapeutic agents,immunotherapeutic agents, chemotherapy, and for blood-brain-barrierdisruption. Methods disclosed herein may include producing bursts ofmonophasic or biphasic pulses for treatment of tumors, alone or incombination with immunotherapeutic agents and chemotherapy. Methods ofelectroporation disclosed herein may include operating an MRI-compatibleelectroporation probe with a grounding pad to deliver pulsed electricfields, either alone or in combination with one or a plurality ofmolecular agents. Such molecular agents may include, for example,chemotherapy, gene-therapy, and tumor-associated antigens. Methods ofelectroporation disclosed herein may provide, for example, reversibleelectroporation for electrogene therapy, electrochemotherapy, andelectroporation to induce immune response. Methods of electroporationdisclosed herein may provide any waveforms capable of inducingelectroporation. Such waveforms may include, for example, reversibleelectroporation for drug delivery, irreversible electroporation (IRE)for tissue destruction, high-frequency irreversible electroporation, andsub-microsecond pulsed electric fields for intracellular manipulation.

In embodiments, methods of electroporation may include number ofelectrodes, size of electrodes, or both, which may vary in relation toshape of a tumor to be treated. Embodiments may include one, two, ormore electrodes, each independently sized and shaped in relation to, anddepending upon, size and shape of a tumor to be treated, size and shapeof killing zones, or both. In embodiments, methods of electroporationmay include varying application of voltage(s) depending upon or inrelation to the size and shape of a tumor to be treated. In embodiments,methods of electroporation may include varying polarity of electrodesdepending upon or in relation to the size and shape of a tumor to betreated, to achieve different electrical fields.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1A is a simplified front view of an MRI-compatible mono-polarelectrode in an exemplary embodiment, with internal structure shown inbroken lines, and with the guide cannula and insulative sheath apartfrom the conductive tubing.

FIG. 1B is a side view of the MRI-compatible mono-polar electrode, takenalong 1B-1B in FIG. 1A, with the guide cannula and insulative sheathapart from the conductive tubing.

FIG. 1C is an isolation top view of a stopper/set screw combination ofthe MRI-compatible mono-polar electrode, taken along 1C-1C in FIG. 1A.

FIG. 1D is a perspective view of the MRI-compatible mono-polarelectrode, taken along 1B-1B in FIG. 1A, with the guide cannula andinsulative sheath joined with conductive tubing.

FIG. 2A is a simplified view of the MRI compatible mono-polar electrodeshown generally in FIG. 1 , assembled with guide cannula andstopper/set-screw combination on.

FIG. 2B is a simplified view of the MRI compatible mono-polar electrodeshown generally in FIG. 2B, unassembled with guide cannula andstopper/set-screw combination off.

FIG. 3A is a simplified perspective assembly view of a cranial boltsystem for mounting to the skull and inserting the electrodes, showingmultiple mono-polar probes in a treatment configuration.

FIG. 3B is a simplified top partial isolation view showing the insertionarray of the electrode and taken generally along 3B-3B in FIG. 3A.

FIG. 3C is a partial isolation front view taken generally along 3C-3C inFIG. 3B, showing the cranial bolt with internal detail shown in brokenlines, cranial bolts. C) Cranial bolt front view.

FIG. 3D is a isolation perspective view showing the cranial bolt of FIG.3B.

FIG. 3E is a partial perspective view of the cranial bolt system shownin FIG. 3A, showing a single-port cranial bolt with ball mechanism toset/adjust insertion angle/trajectory.

FIG. 4A shows a partial perspective view of a full port probe tip, showngenerally in FIG. 3A, in an embodiment.

FIG. 4B shows a partial perspective view of a step-down port probe tip,shown generally in FIG. 3A, in an embodiment.

FIG. 4C shows a partial perspective view of a fillet plug probe tip,shown generally in FIG. 3A, in an embodiment.

FIG. 4D shows a partial perspective view of a chamfer plug probe tip,shown generally in FIG. 3A, in an embodiment.

FIG. 4E shows a partial perspective view of a leaflet valve probe tip,shown generally in FIG. 3A, in an embodiment.

FIG. 4F shows a partial perspective view of an umbrella valve probe tip,shown generally in FIG. 3A, in an embodiment.

FIG. 5A is a schematic diagram showing an electroporation treatment inan exemplary embodiment, with two mono-polar probes inserted into braintissue through guide cannulas.

FIG. 5B is a line graph showing waveforms and pulse parameters forIntegrated Nanosecond Pulsed electroporation treatment method in anexemplary embodiment corresponding to FIG. 5A.

FIG. 6 is a schematic diagram of an electroporation treatment in an MRIsuite, in an exemplary embodiment.

FIG. 7 is a collection of images from MRI compatibility testing forartifact/noise reduction in a method of electroporation, in anembodiment.

FIG. 8 is a functional diagram showing an electroporation systemincluding a pulse generator, shielding port, MRI scanner, and electronicswitch for separating cabling into multiple segments, in an exemplaryembodiment.

FIG. 9A is an image of treatment on a potato using an IntegratedNanosecond Pulsed electroporation treatment method with a 2-5-2 μspulsing sequence, showing an MRI artifact left by nickel titaniumelectrodes disconnected from the generator before treatment.

FIG. 9B is an image of treatment on a potato using an IntegratedNanosecond Pulsed electroporation treatment method with a 2-5-2 μspulsing sequence, showing an MRI artifact left by nickel titaniumelectrodes connected to the pulse generator.

FIG. 9C is an image of treatment on a potato using an IntegratedNanosecond Pulsed electroporation treatment method with a 2-5-2 μspulsing sequence, showing the ablation zone of a 200-burst treatment,seen as the ovoid, contrast-enhancing area surrounding the nickeltitanium electrodes.

FIG. 10 shows Method 100, which is illustrated by a flow chart of aprotocol for electroporation treatment of tissue with simultaneousoperation of an MRI scanner. The steps described herein represent thosethat would follow an initial surgery for placing a cranial bolt andcannula.

FIG. 11A is a Field Emission Scanning Electron Microscopy (FESEM) imageof nitinol, in an exemplary embodiment.

FIG. 11B is a Field Emission Scanning Electron Microscopy (FESEM) imageof nitinol, in an exemplary embodiment.

FIG. 11C is a Field Emission Scanning Electron Microscopy (FESEM) imageof nitinol, in an exemplary embodiment.

FIG. 11D is a Field Emission Scanning Electron Microscopy (FESEM) imageof nitinol, in an exemplary embodiment.

FIG. 11E is a Field Emission Scanning Electron Microscopy (FESEM) imageof nitinol, in an exemplary embodiment.

DETAILED DESCRIPTION

Detailed embodiments of the present disclosure are herein. It may beunderstood that the disclosed embodiments are merely examples toillustrate aspects that may be embodied in various and alternativeforms. Well-known materials or methods have been described in detail toavoid obscuring the present disclosure. Therefore, specific structuraland functional details disclosed herein are not to be interpreted aslimiting, but as a basis for the claims and for teaching one skilled inthe art to variously employ the present disclosure.

This disclosure describes new devices and procedures to improve patientsafety and treatment outcome for patients treated with electroporationtherapy. The methods described are applicable for all ailments withinthe brain, head, neck, spinal cord or other specific locations withinthe body that may be treated with electroporation, including but notlimited to tumors, essential tremors, and Parkinson's disease. The newprocedures described apply to all waveforms capable of inducingelectroporation, including reversible electroporation for drug delivery,irreversible electroporation (IRE) for tissue destruction, integratednanosecond pulse irreversible electroporation, and sub-electroporationeffects for blood-brain-barrier opening. The present disclosure reducesthe risks and enhances treatments of patients receiving electroporationbased procedures.

Referring to FIG. 1 , which contains a set of engineering drawings ofthe MRI compatible mono-polar probe (i.e., electrode) (1) withaccessories, including a guide cannula (5) and stopper/set-screwcombination (4). The mono-polar probe consists of an insulative handle(1), adjustable insulative sheath (2) to control the location ofelectroporation, and conductive tubing (3) to deliver theelectroporation pulses. The conductive tubing (3) extends into thecenter of the handle (1) to permit connection with electrical wiring(i.e., cable; not shown). The connection may be made with a removable,pressure fit connector, or joined permanently. The guide cannula (5)also contains an insulative sheath (6) through which the electrode isinserted. The guide cannula (5) or electrode (1) can also be utilized todeliver exogenous agents or place sensors for monitoring treatment. A)mono-polar probe (1) front view with guide cannula (5) andstopper/set-screw (4). B) Mono-polar probe side view. C) Mono-polarprobe bottom view. D) Mono-polar projection.

FIG. 2 is a set of pictures of the fabricated MRI compatible mono-polarprobe. A) Assembled with guide cannula (5) and stopper/set-screw (4) on.B) Unassembled with guide cannula (5) and stopper/set-screw (4) off.

FIG. 3 is a set of engineering drawings and a picture of a cranial boltsystem (7) for mounting to the skull and inserting the electrodes. A)Assembly view showing multiple mono-polar probes (1) in a treatmentconfiguration. B) Crain bolt top view with insertion array. C) Cranialbolt front view. D) Fabricated cranial bolt. E) Single-port cranial boltwith ball mechanism to set/adjust insertion angle/trajectory.

Referring to FIG. 4 , which contains a set of engineering drawings fordifferent probe tip configurations. Port or valved configurations aredesigned to permit the delivery of exogenous agents via fluid infusion(e.g., chemotherapy), fluid retrieval from the tissue (e.g.,micro-dialysis), or measurement of tissue properties (e.g., electrical,thermal, mechanical) via the placement of sensors. A) Full-sized portfrom hollow conductive tubing (3). B) Step-down port using insulativeinsert. C) Fillet-plug using insulative insert. D) Chamfer-plug usinginsulative insert. E) Leaflet valve using insulative cap. F) Umbrellavalve using insulative cap. An embodiment could also include astandalone cap without a valve or port mechanism.

Referring to FIG. 5 , which is a schematic diagram of a typicalelectroporation treatment. A) Two mono-polar probes are inserted intobrain tissue through guide cannulas (5). All tissue exposed to anelectric field>500 V/cm is subject to electroporation. B) Typicalwaveform and pulse parameters for integrated nanosecond pulseirreversible electroporation treatment.

Referring to FIG. 6 , which is a schematic diagram of an electroporationtreatment in an MRI suite. The electrodes are attached to cables thattravel through a shielded port to the pulse generator, which is housedin the control room. There are multiple points (8) along the length ofthe cabling where it may be disconnected/switched-open and separatedinto multiple segments. This improves safety by reducing the complexityof cabling during treatment preparation (e.g., patient transport) andreduces imaging artifacts/noise propagation when disconnected atdifferent points.

Referring to FIG. 7 , which is a collection of images from MRIcompatibility testing for artifact/noise reduction. Each image shows twomonopolar electrodes with different segments of cabling connected.Descriptions of each scenario are listed below the image.

Referring to FIG. 8 , which is a functional diagram of the pulsegenerator, shielding port, and MRI scanner. A representation of anelectronic switch is shown to indicate one possible method of separatingthe cabling into multiple segments. This could also be accomplished byelectrical plugs, and the location can vary anywhere along the length ofthe cabling, including the electrode handle.

Referring to FIG. 9 , which is a collection of images showing before andafter images of treatments on potatoes using an integrated nanosecondpulse irreversible electroporation pulsing sequence (2-5-2 μs). The MRIartifact left by the Nickel Titanium electrodes when they are (a)disconnected from the generator before treatment and (b) connected tothe generator. (c) The ablation zone of the 200-burst treatment, seen asthe ovoid, contrast-enhancing area surrounding the electrodes.

Referring to FIG. 10 , Method 100 is described by a flow chart of anexample protocol for treating tissue within an MRI scanner. An exemplaryembodiment would include the surgical steps required to gain access tothe brain via a craniectomy or burr hole. An elective pathway isprovided to unplug the cables (mechanically or electrically) in betweensets of pulses to reduce image artifacts/noise. Electrodes are placed inthe tissue (9) based on the tumor geometry to ensure adequate coverageof the tumor with a threshold electric field. This could be accomplishedwith a stereotactic frame and/or infrared neuronavigation system, inwhich infrared tracking balls may be placed on the electrode handledirectly, or as an attachment. Placing (9) may also include mounting acannula (5) to a cranial bolt (7). The insertion depth is controlled byplacing the stopper/set-screw (4) on the electrode shaft. The insertiondepth and electrode position could also be controlled by operating amechanical element. The electrode placement is validated with theinfrared tracking system and/or medical imaging (10). This may occursimultaneously or sequentially (as shown) with electrode placement (9).If the placement is correct (11) a baseline image is collected (13) toestablish pre-treatment contrast and/or temperature; if the placement isnot correct the electrode placement is adjusted (12). The cables areconnected to the electrodes (e.g., at the handle) (14), and baselinetissue properties are calculated (15). A low-voltage test pulse isapplied (16) to ensure proper electric contact between the electrodesand the tissue, and to ensure that, if the voltage were scaled-up to thetreatment level then, according to Ohm's law, the treatment protocolwould fall within an acceptable generator current limit or safety limit(17). If the current is within an appropriate range, the voltage isincreased to the treatment level and the treatment is applied (19). Ifthe current is too low/high, the pulse parameters may be adjusted (18)and/or the electrodes may be re-positioned (12). Operating the pulsegenerator for applying the treatment involves entering the electrodespacing, calculating the applied voltage necessary to achieve a specificvoltage-to-distance ratio, programming the pulse protocol (e.g., pulseduration, number, and repetition rate). The pulse generator couldtrigger the MRI scanner to acquire an image between pulses, or the MRIscanner could trigger the pulse generator to deliver pulses betweenimage acquisitions. During treatment, there is the option to unplug thecables between pulses or groups of pulses (21), acquire an image (22),and re-plug the cables (23). There is also the option to leave thecables plugged-in and make measurements of tissue properties (20) oracquire images. Following treatment, the ablation is verified againstsome metric (24) and the procedure either ends (25) or starts over againfor a new set of electrodes in a different portion of the tumor (1).

Referring to FIG. 11 , which is a series of Field Emission ScanningElectron Microscopy (FESEM) images of nitinol. The first image (11A) issimply of nitinol under no pulse configurations and without insertioninto Phosphate Buffer Solution (PBS, purchased from AmericanBioInnovations, Sparks, Md., USA). Image 11B is of nitinol under nopulse configurations and inserted into PBS. All other images (11C-E)were pulsed at 90 pulses and 70 μs with different voltages in PBS; image11B with 1000V, 11C with 1500V, and image 11D with 2000V.

In embodiments, an Integrated Nanosecond Pulsed IRE system may providemore predictable ablations than systems providing longer waveforms andnon-bipolar waveforms. In embodiments, short bi-polar waveforms providedby an Integrated Nanosecond Pulsed IRE system may provide ablations ofimproved predictability, such as by creating an electrically homogenousenvironment via capacitive coupling. Predictability may be of benefit inthe ablation of portions of the brain with a predominance of whitematter, which are anisotropic and have a higher conductivity along thelong axis of the fibers versus perpendicular to the fibers. An exemplarylist of advantages for Integrated Nanosecond Pulsed IRE systems,including reduced procedural time, reduced procedural complexity, andvisualization of electroporation treatment in real-time under MRIguidance, is provided in Table 1. In embodiments, an IntegratedNanosecond Pulsed IRE system may be an H-FIRE™ Integrated NanosecondPulsed IRE system available from the present applicant (Voltmed Inc.,Blacksburg, Va.).

TABLE 1 Clinical features, advantages, and direct patient benefits ofusing Integrated Nanosecond Pulsed IRE system TECHNICAL CLINICAL FEATUREADVANTAGE BENEFIT Selective killing Ablation of cancer cells Decreaseslikelihood of cancer cells beyond tumor margin of tumor recurrenceNon-thermal method Can treat tumors in Treatment of inoper- of celldeath with proximity to major able tumors and fast electric fieldsvessels/nerves ablation recovery Non-thermal energy Faster than otherReduced procedure delivery completed treatment time and surgical inminutes complications Higher threshold for Does not require generalEnables non-thermal electrically induced anesthesia or paralytics IRE inhighly muscle stimulation sensitive areas or awake surgery More uniformelectrical Penetrates through Generates more properties across differ-tissue/tumor predictable ablations ent tissue types and heterogeneitiesin tissue/tumor post-electroporation MRI-compatibility probe Providesphysicians with Complete tumor for real-time feedback visualization oftreatment coverage improves therapy success

The present disclosure provides improved Integrated Nanosecond PulsedIRE treatment therapy. Embodiments may include a cannula-based electrodesystem. This may include monopolar MRI compatible electrodes. Suchelectrodes may be constructed with a rod approximately 10 cm long thatis coated with a metal, such as copper, on one end so that the metal maybe soldered to the wire connection. Alternatively, the metal may beuncoated and act itself as the “male” end to a paired “female” connector(e.g., banana jack). Nickel titanium may be used as the electrodematerial due to its small MRI artifact in an embodiment, but othermetals, such as platinum, iridium, carbon and silicon may also be used.To visualize the electrode during placement, a ring or an insertcomposed of an MRI contrast material such as Gadolinium may be includednear the tip of the electrode. In some embodiments, the ring may notexceed more than about 1 mm, which is similar to the resolution limit ofMRI. This ring may be used as a bright landmark on MRI imaging tovisualize when the probe has reached its target location, ensuringcorrect depth of insertion and correct spacing. In addition toGadolinium, other material that enhance the contrast of MRI may be usedsuch as manganese, iron oxide, and iron platinum. After the electroderod is plated and soldered, or plugged-in to the wire, a handle, whichhas been custom designed using CAD software, is placed around the solderjoint, or bare metal, and sealed using an autoclavable material such asepoxy (Epoxy International, FDA-Bond 2) and a super glue, such asLoctite 4541, which are both autoclavable materials. Finally, a piece ofinsulation, such as MicroLumen, code475-V.5, Lot #24178, code560-IV, Lot#41614; 8 cm is cut and a stopper, which screws or locks into the skullor a pre-existing skull mount is added to one end of the insulation andsealed with the same super glue. The MRI compatible electrodes may allowfor the electrodes to be guided more precisely and the extent ofelectroporation to be directly observed by MR imaging. The treatment maythen be monitored and changed as needed with MRI guidance.

For purposes of treating brain tumors, the probes to be used in thesystem may consist of monopolar or bipolar electrodes approximately18-gauge, catheter electrodes, and a dispersive electrode (groundingpad), all MRI-compatible and single-use. The tip may be, as described inFIG. 4 , open, port, valved, or plugged/capped. The number andarrangement of electrodes may vary. In one embodiment, no less than 2(including single bipolar probe with an energized and ground surface),and no more than 6, probes may be active simultaneously and may belimited based on current and voltage capabilities. The probes may beidentified by the system through RFID tagging to ensure disposal afterfirst use in clinical settings.

The integrated nanosecond pulse irreversible electroporation system, inembodiments, may include a pulse generator, a monopolar probe, a bipolarprobe, a catheter probe, and a dispersive electrode. The pulse generatormay operate outside the sterile field and in some cases in an MRIcontrol room (FIG. 6 ). The principal components of the generator (FIG.8 ) consist of power electronics, a touchscreen display, and double footswitch pedal, all of which are housed in a wheeled cart. The end usermay interact with the electronics through a graphical user interface onthe touchscreen. Voltage, amperage, and impedance across activeelectrodes may be displayed throughout procedures. The generator mayconnect with a set of MRI-compatible probes through 6 high-voltageoutput connectors. The output parameters consist of high-frequencybiphasic pulses with a maximum voltage and amperage of 5000 Volts and100 Amps, respectively. In clinical settings, the output may be enabledby three possible triggers: signal from a double foot switch pedaloperated by end user, signal from cardiac synchronization device, andsignal from MRI unit to sync with image acquisition sequence. The outputcan also be disabled by either the foot switch or patient-triggeredmanual shut off. The generator may have analog and software safetycomponents that may detect any system malfunctions includingmisconnections, over-currents, and over-voltages.

A method and system of electroporation may include MRI compatiblematerials. Such a system may include a guide cannula for preciselocation and placement of electrodes. Methods for electroporation mayinclude connecting and disconnecting the cabling along its length atmultiple points. An electroporation system may include differentelectrode tip embodiments for different functions (e.g., treatment only,fluid delivery, treatment monitoring).

An electroporation system may include a cannula-based electrode system,with a guide cannula for placing electrodes. This may allow forelectrodes to be strategically placed to minimize distortion fromheterogeneities or maximize ablation within the tissue. The guidecannula may be secured inside a cranial bolt with an array of guidanceholes (FIG. 3A) that is mounted to the skull. An embodiment may includea cranial bolt designed for a single electrode with a tunable insertionangle (FIG. 3E). This may be accomplished through a ball and set-screwcombination. The guide cannula may be imaged with MRI to ensure theproper trajectory before inserting the electrodes. The guide cannula mayenable more accurate, precise placement of the electrodes in tissue andmay provide more accurate treatment area.

The method for placing electrodes may strategically involve the use ofmedical imaging to make a numerical model of the treatment volume. Thismodel may incorporate the tissue geometry via image acquisition andelectrical properties of various tissue types or heterogeneities. Next,an optimization algorithm is developed for placing the electrodes, suchthat the predicted ablation volume resembles a theoretical solution forhomogeneous tissue and/or is the largest obtainable ablation. Forexample, the target tissue may be exposed to an electric field>=500 V/cm(FIG. 5A). To improve imaging, low current may be applied to the tissueduring image acquisition to counteract magnetic field heterogeneitiesand image artifacts caused by the presence of the electrode.

The cannula may be endowed with a variety of properties to improve itsefficacy. The cannula may act as insulation. The cannula may be made ofMRI compatible materials to allow for observed procedures. Usefulmaterials for the cannula include polyimide, Teflon, or PEEK. Anotherembodiment includes mounting the cannula and/or electrode to astereotactic frame without the use of a cranial bolt. Additionally, arobot arm may be used to place the cannula/electrode and electrode totightly control insertion depth. This robot arm may be linked to animage acquisition system where the desired treatment zone may be drawnby the physician and visualized using contrast.

In embodiments, an integrated nanosecond pulse irreversibleelectroporation system may include a Pulse Generator System. Such apulse generator system may be delivered through a custom-builtintegrated nanosecond pulse irreversible electroporation waveformgenerator (VoltMed Inc., Blacksburg, Va., USA) coupled with two (2) ormore blunt-tip electrodes (Ø=1.2 mm; 200-104302; Angiodynamics, Inc.Queensbury, N.Y., USA). The electrodes may have an overall length of 15cm, and may be connected to the pulse generator via 1.8 m insulatedcables. The active exposure length of the electrode tips may be adjustedin 5 mm increments over a range of 40 mm.

The electrodes may be further improved in a variety of ways. Typically,in IRE procedures, non-shielded, insulative electrode leads are used.The disclosure described may add shielding to the electrode leads. Thismethod may help reduce radiofrequency noise and image artifacts, whichmay permit more precise control of the ablation area. Additionally,electrodes may be improved by acting as both the ablation device and thephysiologic sensor. Electrodes may be equipped for real-time measurementof tissue impedance using sensor electrode array placed along aninsulative sheath. These modified electrodes may be used in combinationfor reversibly or irreversibly electroporating tissue and monitoringelectrical activity. The electrophysiological data gathered by themonitoring electrodes may be incorporated in treatment planning forelectroporation procedures.

In IRE procedures, an important adverse effect of treatment is thermaldamage. The disclosure may minimize unwanted thermal damage by directlyobserving the temperature increase as a result of treatment. This isdone by using a temperature probe on the electrode. With activeobservation of the temperature, pulse duration and/or duty cycle may beadjusted during treatment to prevent temperatures lethal to cells frombeing reached. Additionally, the disclosure can utilize a method forperforming MRI thermometry and adjusting the pulse duration and/or dutycycle to prevent lethal temperatures from being reached.

Another adverse effect commonly encountered in electroporation therapyis the interaction of the pulses with critical structures. The presentdisclosure utilizes a method of directing current away from criticalstructures, such as ventricles. This involves placing insulativematerial along the electrode that is positioned between the conductivesurface and the structure to be spared. An external grounding pad orarray of electrodes may be used to direct the current away from thecritical structure. The ventricle or cerebral spinal fluid surroundingthe brain can also be used as a “fluid electrode” by inserting aconductive probe into those regions. This allows for larger treatmentvolumes and more diffuse regions of electroporation. A grounding pad maybe used in combination with needle electrodes in this system and may bemodified in a variety of ways in improve outcome. The needle electrodesand grounding pad may be used so that the bulk tissue electricalresistances local to the electrodes and grounding pad are modified tomaximize the area of treatment. Additionally, all electrodes involved inthe ablation area may be cooled down prior to insertion to minimizeincreases in tissue conductivity near needle-electrodes. A fluid of lowconductivity may be delivered in the vicinity of the needle-electrode toprevent increases in electric field magnitude near the dispersiveelectrode. This may result in larger ablations near the needle-electrodeand minimizing muscle stimulation.

Neuronal networks can also be adversely affected by treatment in thebrain. The disclosure provides a method for controlling pulse timing tominimize effect on neuronal networks. This method consists of using atrain of high-frequency, biphasic pulses, in which the phase delaybetween pulses of alternating polarity is controlled to minimize ormaximize nerve desynchronization. This could also involve purposelyadministering electrical stimulation to create a latency period inneuronal activity, during which electroporation pulses are administered.The method may be improved by synchronizing the delivery ofelectroporation pulses before, during, or after the recording ofinherent or evoked electric potentials.

It is understood that the electrodes could be expanded or reduced inboth length and circumference to allow for larger ablation zones withinother areas of the body. The conductive material used for electrodes maybe a semi-conductive material. This semiconductor may be, but is notlimited to, carbon or silicon-based materials.

An autoclavable material such as Loctite 4541 and Epoxy International,FDA-Bond 2, are used for the construction of the probes in an embodimentbecause they are both autoclavable materials as well as are FDAapproved. However, other FDA approved autoclavable materials may also beused. The electrode may be made of a metal that has small MRI artifacts,such as nickel titanium. The pulse parameters for integrated nanosecondpulse irreversible electroporation (FIG. 5B) are different from otherIRE pulses. They are shorter and bipolar, which is essential forcreating a uniform current distribution, preventing electrically-inducedmuscle contractions, and limiting electrochemical effects. The exposed,non-insulated portion of the electrode controls where the therapy isdelivered. No therapy would be delivered if the guide cannula werelonger, or the same length as, the electrodes. Another embodimentincludes an adjustable length cannula or region of electrode insulation,such that the extent of treatment may be modified, or the electrode maybe covered following treatment but before retraction.

The system described can utilize switching mechanisms, as seen in FIGS.6 and 8 , that may be in the MRI room near the feed through to thecontrol room, and near the elegy rode handle, directly before theelectrodes themselves. This switching mechanism may be a physicaldisconnect or an electrical disconnect. The electrical disconnect couldinclude but is not limited to a physical switch, relay switch, and orsoftware switch. The electrical or physical switching mechanism may becontrolled by human or robotic systems. Disconnecting the cabling nearthe electrode handle reduces noise and image artifacts (FIG. 7 ).

According to the present disclosure, embodiments may combine MRIcompatible electrodes to administer the therapy, shielding the electrodeleads to prevent emanating radiofrequency noise or control the directionof treatment, disconnecting the cabling at specified locations to limitnoise and image artifacts, placing electrodes strategically within aguide cannula to minimize distortion from heterogeneities or maximizeablation within the tissue, utilizing conductive fluids, innate orexternal, such as cerebral spinal fluid or grounding pads to provide apathway for current return, utilizing different electrode tips fortreatment, monitoring, and fluid infusion, and for timing of theelectrical waveforms with inherent brain electrical activity is toenhance safety and treatment outcomes for patients undergoingelectroporation procedures.

The utility of the system and method disclosed herein has beenestablished by studies. The following examples are meant only to beillustrative and are not meant as limitations on the scope of theinvention or of the appended claims.

The applicant conducted both theoretical and experimental studiescharacterizing the effects of IRE for soft tissue ablation in the brainin a pilot safety and efficacy study on six normal dogs. The resultsshowed that the treatment was well-tolerated and spared major brainvasculature, suggesting that IRE is suitable for treatment of tumorsadjacent to, or enveloping, critical vascular structures. Seven (7)canine patients with aggressive gliomas were successfully treatedwithout causing collateral damage to the surrounding normal brain usingIRE.

Post-IRE magnetic resonance imaging (MRI), histopathology, and clinicalneurological examinations indicated that tumor reduction was achieved insix (6) of the seven (7) canines. The median 14-day post-IRE KarnofskyPerformance Score (KPS) of the 6/7 dogs surviving to discharge was 80(range 60-90), and was improved over pre-treatment values in all cases.Objective tumor responses were seen on MRI in ⅘ of dogs withquantifiable target lesions. Two dogs experienced survivals in excess ofone year, including 1 dog that demonstrated a complete response to IREtreatment for 4+ years to date. With other therapies, median survivalsof 0.2, 0.9, and 4.9 months have been reported for dogs with braintumors receiving either symptomatic therapy, cytoreductive surgery, ormultimodal therapy (surgery and radiotherapy or hyperthermia),respectively, illustrating the poor prognosis.

A technical challenge at the time was the strong muscle contractionsinduced by IRE pulses. This required the use of enhanced anestheticprotocols that included neuroparalytic agents. Even then, pulse inducedmovement was still present and shown to disrupt electrode-tissuecontact. Dislodging of the electrodes poses a risk for the more than 50%of patients with brain tumors in poor surgical locations (e.g., corpuscallosum, motor cortex, insula, multiple locations, etc.). Additionally,muscle paralysis involves general anesthesia and mechanical ventilation,which precludes the use of IRE in ‘awake’ neurosurgical interventions orin severely debilitated patients.

A prospective, single center, pilot study of canine patients wasdesigned according to IDEAL stages 1/2a of surgical innovation toevaluate the feasibility and safety of ablating brain tumors with.Client-owned dogs with naturally occurring intracranial meningiomas wererecruited through the treatment center's referral network and byregistry of the trial on a national veterinary clinical trials database.To be eligible for the trial, dogs had to have clinical signs of braindisease, a diagnostic brain magnetic resonance imaging (MRI) scandemonstrating a solitary mass lesion>1 cm in diameter with imagingcharacteristics compatible with a meningioma, Karnofsky PerformanceScore (KPS)≥60, and be free of significant concurrent cardiopulmonary,renal, and hepatic disease, or other malignancy. When applicable, dogswith structural epilepsy had to have seizures that were controlled onanticonvulsant medications. Exclusion criteria included the receipt ofany type of prior brain radiotherapy, or treatment with a cytotoxicchemotherapy drug within 6 weeks of trial enrollment. Owners providedwritten, informed consent to enroll their dogs into the study.

Testing of the MRI compatible probes were conducted and shown in FIG. 7. The artifact, as seen from these images, shows it being only 5 mm froma 1 mm diameter probe. Each image also shows how the image is affectedby the connection of the electrode to each specific connection point asillustrated in FIG. 6 . Also imaged was the guide cannula alone in thecopper sulfate (CuSO4) solution, which may be seen in FIG. 7 as theimage on the bottom row and furthest on the right. The series of imageswere taken with a spin echo with the following specifications: TR—500.0ms, TE—2.7 ms, and BW— 930.0 Hz. The images were imaged with the use ofa 1.5 MRI scanner (Phillips Medical Systems, Winston-Salem, N.C., USA).

FESEM was used to image nitinol (purchased from VascoTube GmbH,Baden-Wuerttemberg, Germany) and are shown in FIG. 11 . Imaging was doneat the ICTAS Nanoscale Characterization and Fabrication Laboratory(Blacksburg, Va., USA) using their LEO 1550 (Zeiss, Dublin, Calif., USA)which is capable of resolution in the 2-5 nm size range. For all of theimaging, we used the SE2 detector for general purpose imaging and at anElectron High Tension (EHT) value of 5 kilovolts; EHT is the voltagethrough which the electrons are accelerated toward the sample. Themagnitude of all the samples was 200×. Immersion testing was done usingPBS due to the similar pH, osmolarity, and ion concentrations as that ofthe human body. The samples were pulsed using a BTX T820 pulse generator(Holliston, Mass., USA) and, using gloves to avoid contamination,mounted onto aluminum stubs, which were fixed onto the ZEISS specimenholder using carbon tape.

As seen from the images, there is no apparent difference in surfacecondition due to pulsing and varying voltages. The crystal structuresseen in FIG. 11B are likely NaCl from the PBS. We know this because thecrystal structures are absent in FIG. 11A, where nitinol was notimmersed in PBS. It appears the presence of cracks and contamination onthe surface stays relatively constant; those in FIGS. 11D and 11E aredue to cutting of the nitinol wire at those points.

Potatoes were treated with a variety of pulse sequences along withdifferent burst amounts to show the ablation sizes that could beachieved. Each potato was imaged before treatment and after treatment.In FIG. 9 , the ablation this testing may be seen with the 2-5-2 pulse(μs) sequence and both the 40-burst along with the 200-burst amount andthe ablation sizes of each. Each post image was taken almost directlyafter the treatment. All images were taken by a 1.5 MRI scanner(Phillips Medical Systems, Winston-Salem, N.C., USA), imaging with aflair dorsal with a TE of 3.4 ms. There is an optimized scan for the 1.5MRI scanner called a UTE (ultrashort echo time) where the TE is loweredto 0.05 ms and this scan can reduce the artifact of the electrodes tojust 2.6 mm.

A prospective, single center, pilot study of canine patients wasdesigned according to IDEAL stages 1/2a of surgical innovation toevaluate the feasibility and safety of ablating brain tumors with.Client-owned dogs with naturally occurring intracranial meningiomas wererecruited through the treatment center's referral network and byregistry of the trial on a national veterinary clinical trials database.To be eligible for the trial, dogs had to have clinical signs of braindisease, a diagnostic brain magnetic resonance imaging (MRI) scandemonstrating a solitary mass lesion>1 cm in diameter with imagingcharacteristics compatible with a meningioma, Karnofsky PerformanceScore (KPS)≥60, and be free of significant concurrent cardiopulmonary,renal, and hepatic disease, or other malignancy. When applicable, dogswith structural epilepsy had to have seizures that were controlled onanticonvulsant medications. Exclusion criteria included the receipt ofany type of prior brain radiotherapy, or treatment with a cytotoxicchemotherapy drug within 6 weeks of trial enrollment. Owners providedwritten, informed consent to enroll their dogs into the study.

On the day of admission (Day 1), dogs underwent pre-treatment KPSscoring, and complete physical, neurological, and laboratoryexaminations. They were anesthetized using a complete intravenousprotocol consisting of premedication with methadone and midazolam,induction with propofol, and maintenance with propofol and remifentanilconstant rate infusions. Anesthetized dogs were instrumented in an MRIcompatible, small animal stereotactic headframe (Dynatech, DynatechMachining, Union City, Calif., USA). MRI images of the brain wereobtained for therapeutic planning (see Supplementary Digital Content1—Methods, MRI Protocol) as previously reported. Parasagittalmeningiomas were classified using the Sandor schema after acquisition ofMRI venograms (very). After stereotactic images were obtained, dogs wererecovered from anesthesia.

Patient-specific integrated nanosecond pulse irreversibleelectroporation treatment plans were developed using MRI-based tissuesegmentation, volumetric meshing, and finite element modeling accordingto previously described methods (see Supplementary Digital Content1—Methods, Treatment Planning). The therapeutic planning procedure wascustomizable and generated three-dimensional patient and tumor specificoutputs. These outputs depicted the expected electric field distributionand Joule heating given the electrode approach and configuration foreach electrode pair being used in the treatment.

On Day 2, dogs were placed under general anesthesia, instrumented in thestereotactic headframe, and aseptically prepared for surgery. To monitorfor muscle contractions, a 3-axis accelerometer breakout board (ADXL335,Adafruit Industries, New York, N.Y., USA) with a sensing range of ±3 gwas sutured to the skin of each dog in the dorsal cervical region at thelevel of the C2 vertebra. In the operating theater, each dog underwent acraniectomy approach of sufficient size to expose the tumor forintegrated nanosecond pulse irreversible electroporation systemtreatment and subsequent tumor resection. Following completion of thetumor exposure, incisional biopsies of the tumor were obtained.Integrated nanosecond pulse irreversible electroporation systemtreatments were delivered stereotactically according to pretreatmentplans by mounting and advancing the electrodes to the target regionusing micromanipulator arms of the headframe. Pulse delivery wassynchronized with the electrocardiogram (Ivy Cardiac Trigger Monitor3000, Branford, Conn., USA) to prevent cardiac arrhythmias. Theelectrodes were removed from the brain, and each patient underwent tumorresection using standard techniques. Following resection, surgicalwounds were closed routinely, and then immediate post-treatment brainMRI examinations performed. All dogs received perioperative antibiotics(cefazolin, 22 mg/kg, IV, q 8 hours) and buprenorphine (0.003 mg/kg, IVor SC, q 6-8 hours) for at least 24 hours following recovery from thehigh-frequency irreversible electroporation treatment. Followinganesthetic recovery on Day 2, and on each subsequent day ofhospitalization until discharge, each dog underwent post-treatment KPSscoring, complete physical, neurological, and laboratory examinations,and adverse event (AE) monitoring. The study ended after each dogcompleted a 14-day post-treatment recheck clinical examination, KPSscore, and AE assessment.

The primary end-point was to evaluate the safety of integratednanosecond pulse irreversible electroporation system for the treatmentof brain tumors. To this study, safety was defined as the absence ofsevere clinical toxicity within 14 days of the integrated nanosecondpulse irreversible electroporation system. Severe toxicity wasclinically defined by a ≥20-point decline in the KPS from pre-treatmentvalues, or development of grades 3, 4, or 5 AE, as classified accordingto the Cancer Therapy Evaluation Program's Common Terminology Criteriafor Adverse Events (NCI CTCAE v4.0).

Secondary end points included direct neurotoxicity evaluationsdetermined from post-treatment imaging studies and morphologicevaluation of tumor ablations. Following resection, each tumor wasimmersion fixed in bloc in 10% neutral buffered formalin for 48 hours.After fixation, the tumor was mounted in matrix slicer (ZeviInstruments, Pittsburgh, Pa., USA), photographed, and then seriallysectioned in the transverse plane at 2 mm intervals. Sections werestained routinely with hematoxylin and eosin (H&E, Sakura Fintech,Torrance, Calif., USA). Light microscopy was used to type and gradetumors according to World Health Organization criteria, and to performqualitative morphometric analyses. Ablation volumes were obtained usingcommercial image analysis software with Cavalieri estimator (StereoInvestigator, MBF Biosciences, Williston, Vt., USA).

Three dogs with intracranial meningiomas enrolled in and completed thestudy between March and July 2016. No evidence of muscle or nerveexcitation or cardiac arrhythmia during any pulse delivery was observedin any dog. Additionally, no displacement was detected by theaccelerometers for any of the delivered pulses, nor were any significantpost-treatment laboratory abnormalities attributable to integratednanosecond pulse irreversible electroporation system detected in anydog.

No intra- or post-operative AE were observed in Dogs 1 and 3, and thesedogs were discharged from the hospital with static clinical examinations24 hours after the integrated nanosecond pulse irreversibleelectroporation system procedure. Intraoperatively, Dog 2 experiencedintracranial hemorrhage and subsequent hypotension following disruptionof a collateral vein during tumor resection. Hemorrhage was controlledwith topical hemostatic agents, temporary venous hemoclipping, and bloodpatches. Post-operatively, Dog 2 developed a depressed level ofconsciousness, an exacerbation of pre-existing hemiparesis, and 10-pointpost-operative decline in KPS score from baseline. Due to intraoperativeAE, immediate post-operative imaging was not performed in Dog 2. Thehypotension resolved upon anesthetic recovery. Dog 2 was discharged fromthe hospital 7 days after the procedure, and its neurological statusreturned to pre-treatment value by the day-14 recheck.

No imaging evidence of direct neurotoxicity or collateral damage tobrain tissues outside of H-FIRE treatment zones was observed in Dogs 1and 3 on immediate post-treatment MR examinations, or in Dogs 1 and 2six months after treatment. In Dogs 1 and 2, ablations completelydisrupted the cytoarchitecture of the tumors, resulting in homogeneousregions of tumor necrosis clearly delineated from adjacent untreatedareas. In Dog 3, ablations resulted in non-uniform treatment regionscharacterized by patchy necrosis and, in areas surrounding psammomabodies, a marked neutrophilic and lymphocytic infiltrate surroundingislands of edematous but viable tumor cells.

Dogs were followed off-protocol for six months or until death. No dogreceived other treatment in the six months following integratednanosecond pulse irreversible electroporation system ablation. Dog 1 wasalive, seizure free, and had no evidence of tumor six months aftertreatment. Dog 2 was alive six months post-treatment, but requiredescalation of anticonvulsant therapy for persistent post-treatmentseizure activity, and had suspected residual or recurrent tumoridentified on MRI examination performed 5 months after treatment. Dog 3died 76 days after treatment due to complications arising from recurrentstatus epilepticus.

An integrated nanosecond pulse irreversible electroporation system andmethod as disclosed herein provides an ablative technique for thetreatment of spontaneous brain tumors. An integrated nanosecond pulseirreversible electroporation system and method as herein disclosed iscapable of safely ablating defined focal areas of normal canine androdent brains as well as spontaneous glioma.

The blood-brain-barrier (BBB) presents a significant obstacle to thedelivery of systemically administered drugs for the treatment of braincancer and neurological disorders, such as Parkinson's disease andessential tremor. According to the present disclosure, two distinctpulse regimes may induce a defined zone of BBB disruption. When ablationprotocols are implemented (1000 V/cm, 90 pulses) a reversible penumbraof BBB disruption may exist outside the zones of complete cell death andpreferential cancer cell death. Additionally, sub-lethal protocols havebeen defined that may utilize a lower applied electric field (250 V/cm)and greater number of pulses (300 pulses). An electric field basedsystem for BBB disruption could represent a clinically useful platform.

Reversible electroporation with a device as herein disclosed may be usedto deliver drugs into cells within the treatment area. Irreversibleelectroporation as disclosed may be used to kill a specific type ofcells within the treatment area. Both may be used together withdifferent ablation zones consisting of each reversible, outer edge, andirreversible electroporation, center of the ablation.

The apparatus and methods described herein may provide treatments forneurological disorders. Such disorders may include, for example, braintumors, essential tumors, and Parkinson's disease. The methods mayinclude pulse waveforms capable of inducing irreversible electroporationfor tumor ablation (with and without accompanying thermal effects),reversible electroporation for drug delivery, and sub-electroporationeffects for blood-brain-barrier opening (e.g., modulation of tightjunctions). Methods as disclosed for controlling pulse timing tominimize effects on neuronal networks may use a train of high-frequency,biphasic pulses, in which the phase delay between pulses of alternatingpolarity is controlled to minimize or maximize nerve desynchronization.In embodiments, the positive phase may be of higher amplitude andshorter duration than the negative phase, such that the overall waveformis charged balanced. In some embodiments, the phases may be of equalduration and amplitude. In embodiments, electrical stimulation maycreate a latency period in neuronal activity, during whichelectroporation pulses are administered. In some embodiments, thedelivery of electroporation pulses may be synchronized before, during orafter the recording of inherent or evoked electric potentials.

The apparatus and method described herein are ways of directing currentaway from critical structures, such as the ventricles. Place insulativematerial along the electrode that is positioned between the conductivesurface and the structure to be spared. Use an external grounding pad orarray of electrodes to direct current away from the critical structure.Treatment electrodes may be placed in the tissue. The side of thetreatment electrode facing the ventricle may be insulated. An externalelectrode array placed in contact with the subject is further used todirect current away from the ventricle. In embodiments, the ventricle orcerebral spinal fluid surrounding the brain can also be used as a “fluidelectrode” by inserting a conductive probe into those regions. This mayhelp create larger treatment volumes or more diffuse regions ofelectroporation.

Systems and methods described herein may enable placing electrodesstrategically to minimize distortion from heterogeneities or maximizeablation. Systems and methods described herein may use medical imagingto make a numerical model of the treatment volume that incorporates theelectric properties of various tissue types or heterogeneities. Systemsand methods described herein may include an optimization algorithm forplacing the electrodes, such that the predicted ablation volume mayresemble a theoretical solution for homogeneous tissue and/or is thelargest obtainable.

Systems and methods described herein may use a previously implanted deepbrain stimulation (DBS) electrode to administer electroporation.Beneficial effects of DBS may become suppressed over time, in which casean ablative or drug delivery method may be utilized to treat the tissue.

The apparatus and method described herein for inserting an electrodethrough a guide cannula may include a cannula which may be mounted toskull following creation of burr hole. In some embodiments, the cannulamay act as insulation. In some embodiments, the electrode and cannulamay be made from MRI compatible materials. In some embodiments, theelectrode spacing may be designed to minimize MRI artifacts. In someembodiments, a low current may be applied to tissue during imageacquisition to counteract magnetic field heterogeneities and imageartifacts caused by the presence of the electrodes. In some embodiments,a robot arm may control insertion depth of cannula and electrode. Insome embodiments, a contrast agent may be used to distinguish treatmentzone from other tissue. In some embodiments, a drawing of a desiredtreatment zone may be prepared.

Systems and methods described herein may shield the electrode leads forcompatibility with operation of MRI equipment. The shielding may extend,for example, the full length of leads to the exposed, energized andgrounded electrodes. Shielding may reduce radiofrequency noise andleakage of current into the tissue outside the treatment site.

Systems and methods described herein may perform MRI thermometry andadjusting the pulse duration and/or duty cycle to prevent lethaltemperatures from being reached. Systems and methods described hereinmay use a temperature probe on the electrode and may include adjustingthe pulse duration and/or duty cycle to prevent lethal temperatures frombeing reached. Systems and methods described herein may includeincreased electrode diameter and may compress surrounding tissue. Inembodiments, systems and methods described herein may apply all of thetechniques above in combination with an adjuvant Chemotherapy,Radiosensitizer, Peptide, or drug Immunostimulatory agent.

The terms and expressions that have been employed are used as terms ofdescription and not of limitation, and there is no intention in the useof such terms and expressions of excluding any equivalents of thefeatures shown and described or portions thereof, but it is recognizedthat various modifications are possible within the scope of theinvention claimed. Thus, it may be understood that although the presentdisclosure has been specifically disclosed by embodiments and optionalfeatures, modification and variation of the concepts, herein disclosed,may be resorted to by those skilled in the art, and that suchmodifications and variations are considered to be within the scope ofthis.

It is understood that embodiments may apply to performing a combinationof electroporation and thermal ablation/damage, such that if the pulseduration or number is increased to allow heating near the electrodes.

Embodiments are within the scope of the disclosure and the followingclaims. In general, the terms and phrases used herein have theirart-recognized meaning, which may be found by reference to standardtexts, journal references and contexts known to those skilled in theart. The preceding definitions are provided to clarify their specificuse in the context of the invention. All references cited herein arehereby incorporated by reference to the extent that there is noinconsistency with the disclosure of this specification. Insofar as thedescription above and the accompanying drawings disclose any additionalsubject matter that is not within the scope of the claims, theinventions are not dedicated to the public and the right to file one ormore applications to claim such additional inventions is reserved.

What is claimed is:
 1. A method for ablating tissue, the methodcomprising: in an imaging room, placing one or more electrode in or neara target region; connecting one or more of the electrodes to a functionpulse generator by way of a cable linkage which passes through ashielded port to a control room separate from the imaging room;operating the function pulse generator to ablate tissue; imaging thetarget region using MRI; wherein a physical and electrical disconnect ofone or more of the electrodes from the cable linkage exists, and whereinthe imaging is performed while one or more of the electrodes ismechanically unplugged from the entirety of the cable linkage to obtaina baseline MRI image before operating the function pulse generator;re-connecting the electrodes by mechanically plugging them into thecable linkage to the function pulse generator; after operating thefunction pulse generator, again mechanically unplugging one or more ofthe electrodes from the entirety of the cable linkage; and after theunplugging, performing a second imaging of the target region using MRI.2. The method of claim 1, wherein: the placing of the one or moreelectrode is performed while one or more of the electrodes are unpluggedfrom the cable linkage.
 3. The method of claim 2, further comprising:after the second imaging, re-connecting one or more of the electrodes tothe cable linkage by plugging the electrodes into the cable linkage;after the re-connecting, operating the function pulse generator a secondtime to ablate tissue.
 4. The method of claim 3, further comprising:repeating the unplugging, second imaging, re-connecting and/or operatingof the function pulse generator until a desired MM image is obtained. 5.The method of claim 1, wherein one or more of the electrodes arecomprised of one or more of the following: electrical conductors or agrounding pad; and/or a counter electrode which is a dispersiveelectrode or ground pad to orient high electric fields in a desireddirection; and/or a needle-electrode and a ground pad; and/or at leastone electrode configured for reversibly or irreversibly electroporatingtissue and monitoring electrical activity.
 6. The method of claim 1,further comprising: creating a burr hole in a skull, optionally underMill guidance; mounting a cranial bolt to the skull and inserting aguide cannula to the skull through the cranial bolt, wherein the guidecannula optionally provides insulation for one or more electrode;inserting one or more of the electrodes into the guide cannula; andinserting a portion of the guide cannula into brain tissue, optionallyby operation of a controllable mechanical device.
 7. The method of claim1, wherein one or more of the electrodes are MRI compatible and arrangedto minimize artifacts and/or noise by using non-magnetic material for aprobe and having MRI compatible electrode connections, with the abilityto reach outside of an MRI suite.
 8. The method of claim 1, wherein oneor more of the electrodes acts as both an ablating electrode and anelectrophysiological sensor.
 9. The method of claim 1, wherein bulktissue electrical resistances local to an electrode and ground pad aremodified by fluid infusion to maximize an area of treatment.
 10. Themethod of claim 1, wherein one or more of the cable linkages compriseshielded leads capable of reducing, limiting or preventingradiofrequency noise, image artifacts or electromagnetic interferencewith the MRI.